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Characterization of Calcium-Activated Bifunctional Peptidase of the Psychrotrophic Bacillus Cereus

Characterization of Calcium-Activated Bifunctional Peptidase of the Psychrotrophic Bacillus Cereus

Jong-Il Kim, Sun-Min Lee, Hyun-Joo Jung

J Microbiol. 2005 Jun;43(3):237-43.

PMID: 15995640

Abstract:

The protease purified from Bacillus cereus JH108 has the function of leucine specific endopeptidase. When measured by hydrolysis of synthetic substrate (N-succinyl-Ala-Ala-Pro-Leu-p-nitroanilide), the enzyme activity exhibited optimal activity at pH 9.0, 60 degrees C. The endopeptidase activity was stimulated by Ca++, Co++, Mn++, Mg++, and Ni++, and was inhibited by metal chelating agents such as EDTA, 1,10-phenanthroline, and EGTA. Addition of serine protease inhibitor, PMSF, resulted in the elimination of the activity. The endopeptidase activity was fully recovered from the inhibition of EDTA by the addition of 1 mM Ca++, and was partially restored by Co++ and Mn++, indicating that the enzyme was stabilized and activated by divalent cations and has a serine residue at the active site. Addition of Ca++ increased the pH and heat stability of endopeptidase activity. These results show that endopeptidase requires calcium ions for activity and/or stability. A Lineweaver-Burk plot analysis indicated that the Km value of endopeptidase is 0.315 mM and Vmax is 0.222 mmol of N-succinyl-Ala-Ala-Pro-Leu-p-nitroanilide per min. Bestatin was shown to act as a competitive inhibitor to the endopeptidase activity.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP70968046	N-Succinyl-Ala-Ala-Pro-Leu p-nitroanilide		70968-04-6	Price

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